In recent years, detections of fecal human hemoglobin (fecal occult blood) caused by bleeding from gastrointestinal tracts have been broadly carried out as a screening method of a digestive system disorder, such as colon cancer. The detective method of human hemoglobin is an immunological method employing a specific antibody against human hemoglobin, which takes the place of a related-art dipstick method based on a chemical coloring reaction. This method does not require a dietary restriction, and therefore it becomes established as a convenient testing method.
Examples of an immunological detective method for human hemoglobin include: a single immunodiffusion test in agar plates employing a precipitation line between an anti-human hemoglobin antibody and a human hemoglobin in a testing sample; a latex agglutination test using a latex particle sensitized with an anti-human hemoglobin antibody; enzyme immunoassay or radioimmunoassay, using an anti-human hemoglobin antibody labeled with an enzyme or a radioactive element; and gold colloid aggregation colorimetric method using a gold colloid particle sensitized with an anti-human hemoglobin antibody.
However, in a testing solution, human hemoglobin is gradually denatured, and its antigenicity is decreased. Also, in a testing solution, storage conditions such as storage temperature often accelerates denaturation of human hemoglobin, or bacteria and digestive enzymes in feces often degrade human hemoglobin. Such denaturation and degradation destroy human hemoglobin conformation, resulting in decrease of antigenicity. Therefore, in an immunological method for measuring human hemoglobin, denaturation and degradation of the hemoglobin involve an incorrect diagnosis.
Meanwhile, on a fecal occult blood test, feces are often collected by subjects themselves at their places, and are provided for the test with a closed container, by dissolving the feces in a feces dissolving solution in the container. In such cases, human hemoglobin in feces is often remained in the solution for several days, or placed in a high temperature when utilizing a transportation method such as a postal service. Also, even when feces are collected in a clinical laboratory, it sometime takes longer until carrying out the fecal occult blood test because other tests are also performed. Under such circumstances, an accurate measurement is disturbed by denaturation and degradation of human hemoglobin as described above.
To prevent such denaturation and degradation of human hemoglobin in a solution, a method of adding common antibacterial agents such as thimerosal and chlorhexidine (see Patent Document 1, for example); a method of adding saccharides (see Patent Document 2, for example); addition of hemoglobin of animals other than human (see Patent Document 3, for example); addition of sera of animals other than human (see Patent Document 4, for example); addition of a bacteriolytic enzyme (see Patent Document 5, for example); addition of iron protoporphyrin (see Patent Document 6, for example) and the like have been developed.
However, the techniques for stabilizing human hemoglobin described in those publications cannot fully suppress denaturation and degradation of human hemoglobin in a testing solution containing feces.
Other than those described above, a method of stabilizing hemoglobin using ethylenediaminetetraacetic acid (hereafter, abbreviated as EDTA) have been developed (see Patent Document 7, for example).
However, as a result of replication studies conducted by the inventors of the present invention, it has been confirmed that a sufficient stabilizing action on fecal human hemoglobin cannot be expected with a single use of EDTA.
Then, the applicant of the present invention has been developing a method of adding an aqueous transition metal complex which is more effective for stabilization than EDTA only (see Patent Document 8, for example). Further, the applicant has already developed a method of stabilizing hemoglobin with which a ferrocyanide compound coexists (see Patent Document 9, for example); a method of stabilizing hemoglobin with which an enzymatic degradation product of hemoglobin coexists (see Patent Document 10, for example); a method of stabilizing a heme protein with which transition metals coexist (see Patent Document 11, for example); a method of stabilizing a heme protein with which an organic acid such as malic acid coexists (see Patent Document 12, for example); and a method of stabilizing a heme protein with which a delipidated albumin coexists (see Patent Document 13, for example).    Patent Document 1: Japanese Patent Application Publication No. JP-A-63-271160    Patent Document 2: Japanese Patent Application Publication No. JP-A-63-243756    Patent Document 3: Japanese Patent Application Publication No. JP-A-2-296149    Patent Document 4: Japanese Patent Application Publication No. JP-A-4-145366    Patent Document 5: Japanese Examined Patent Application Publication No. JP-A-5-69466    Patent Document 6: Japanese Patent Application Publication No. JP-A-5-281227    Patent Document 7: Japanese Patent Application Publication No. JP-A-5-99923    Patent Document 8: Japanese Patent Application Publication No. JP-A-7-229902    Patent Document 9: Japanese Patent Application Publication No. JP-A-11-118806    Patent Document 10: Japanese Patent Application Publication No. JP-A-11-218533    Patent Document 11: Japanese Patent Application Publication No. JP-A-2001-249132    Patent Document 12: Japanese Patent Application Publication No. JP-A-2003-014768    Patent Document 13: Japanese Patent Application Publication No. JP-A-2003-194825